Process for the manufacture of aminoacyl compounds



States Patent c 2,788,341

Patented Apr. 9, 1957 The said S-aminoacylmercapto compounds are new. 2,788,341 They can be prepared by the process described in Appli- PROCESS FOR THE MANUFACTURE OF cation Ser. No. 391,738, filed November 12, 1953. In AMl OACYL C M OUNDS general the process comprises reacting a. compound of 5 the formula AXB1 with. a compound of the formula Robert schwyzerr Rlehen swltzefland to Bz--YZ wherein B1 and B2 represent substituents Pharmaceufical Products summit which react to form the grouping -CO-S linking No Drawing, Application N b 12, 1953, X and Y; and A, X, Y and Z, have the aforementioned Serial No. 391,739 significance.

10 For the reaction with the acylaminomercaptans and Claims pnonty, application Switzerland March 6, 1 their salts, any desired compound containing active hydro- 10 Claims, (Cl. 260-412) gen can be used, e. g. such as contain a hydroxyl or amino group, for instance aliphatic, alicyclic, aromatic,

This invention relates to the manufacture of aminoacyl "irahphatlc or heterocych? amlpes or h in compounds and more particularly to a new process for 5 t cular natural or synthetic arn1nocarboxyl1c acids, pepthe aminoacylation of compounds containing active hymics or protems,

dmgen, as for example, amines or alcolmm The process 1s carried out either without any solvent The introduction of aminoacyl radicals into amino caror. Preferably m presence of l or 9 solvents boxylic acids and estfis thfimof by reacting them with such as alcohols, dimethyl formamule or mixtures thereacylaminocarbothiolic acids and their phenyl esters is Concentrations are advailtageolisly 1 mol or more known Cronyn L Ii, Journal of the Americal of each of the reactants. It 1s possible to employ mild Chemical Society 74 21726 [1952]. J. a Sheehan and conditions, which is of considerable importance in the D. A. Johnson, Journal of the American Chemical Synthesis of .relatively 9 nT i Society, 74, 4726 [1952]; Wieland and coworkers ent process gives good yields, especially when 1t 18 carried Liebigs Annalen der Chemie, 573, 99 [19511; 576, 104 at a PH [1952]; zietschriftrfur angewandte Chemie, 63 146 The followmg examples lllustrate the 1nvent1on, the [1951]). However, the procedure involves a number of Parts. beulg by Welght unliss Otherwlse stated and i drawbacks. For one thing, the acylaminocarbothiolic wlamonshlp of Pans by welght to Parts. by vqlume being acids are relatively difiicult to prepare and are of low the same as that of the gram to the wine centimeter:

stability. Their phenyl esters are sparingly soluble in 39 Example 1 water, so that the reaction has to be carried out in an 0.25 part of S-(benzoylamino-acetyl)-mercaptoacetic organic solvent. This circumstance works as a handicap acid (prepared according to Example 1 in application in the acylation of amino acids, peptides, and proteins, Ser. No. 391,738, filed November 12, 1953, is dissolved which are diflicultly soluble in organic solvents. Morein 2 parts by volume of dimethyl formamide and mixed over, the yields obtained are unsatisfactory. with 4 parts by volume of a citratephosphate-borate The present invention now provides an advantageous buffer of the desired pH (according to T. Theorell and process of aminoacylation y reacting o po s o E. Stenhagen, Biochemische Zeitschrift, 299, 416 [1938]). wining active hydrogen with y pta o After an addition of 0.09 part of aniline, the solution is the formula given the desired pH by means of caustic soda solution A-X'C,OSY-Z 40 or hydrochloric acid. Finally, water is added to make up 12 parts by volume, in other words, to bring the concentration of the reactants to about 0.083 mol. The mixture is allowed to stand at 35 C. for 24 hours at the end of which period the benzoylaminoacetanilide formed has separated in pure form. It melts at 217 C. the crystals are filtered and Weighed.

containing electrophilic substituents, or salts thereof. In

this formula, A represents an amino group which may be free or substituted, e. g. by acyl, acylaminoacyl, aminoacyl aminoacyl, alkyl, alkylene, cycloalkyl, aryl, aralkyl,

and/or heterocyclie radicals, for instance the acetyl-,

aminoacretyl" ammoacetylammoapetyl" carpobenzoxy' The following yields are obtained with the application amino acetyl-, carbobenzoxyarmnoacetylannno-acetyl-, of the Stated PH values.

benzoyl-, phthaloy1-, nitrophenoxyacetyl-, carbobenzoxy-,

thiocarbobenzoxy-,dhnethylor diethylamino or piperidino PH: Pew?!t or morpholino group. X and Y stand for any organic 2 members which may form isocyclic or heterocyclic rings, 4 66 in particular members wherein A is separated from C0, 2 43 or S from Z, by 1 to 4 carbon atoms, such as 37 the methylene, ethylene, propylene or phenylene 21 group. A-X-CO preferably represents the radical of a natural amino acid, such as an aminoacetyl or amino, The reaction can be illustrated by the following equapropionyl radical. Z stands for an electrophilic subtion:

stituent, especially for the carboxyl group, and also for Example 2 the sulfo, the nitro or an acylamino group or a free, 0.25 part of S-(benzoylamino-acetyl)-merapto-acetic ,etherified or esterified hydroxyl group or for a halogen acid are reacted in the manner described in Example 1 atom. with 0.11 part of benzylamine according to the equation 3 4 The yield of benzoylaminoacetic acid benzylamide (melt- Example 6 ing at 164 C.) is 43 percent at pH 8 or 73 percent at 034 part of S (carbobenzoxyamino acety1) Thio PH salicylic acid (prepared according to Example 2 in appli- Example 3 cation Ser. No. 391,738, filed November 12, 1953 and 0075 part of amino-acetic acid are reacted at 80-85 C. exactly as described in Example forthe'corresponding mercaptoacetic acid derivative. After working up in the same Way, there is a yield of 0.130 part (=49%) of Ncarbobenzoxyaminoacetyl-aminoacetic acid.

0.25 part of S-(benzoylamino-acetyl)-mercapto-acetic acid and 0.075 part of aminoacetic acid are mixed with 1 part by volume of 2 N-caustic soda solution, and the solution kept at 35 C. for 15 hours. After the addition of 1 part by volume of 2 N-hydrochloric acid, the re- 10 action mass is evaporated to dryness and unchanged S-(benzoylamino-acetyl)-mercapto-acetic acid, and also benzoylamino-acetic acid (formed by hydrolysis), dis- COOH solved in absolute alcohol. The insoluble residue is I Washed with alcohol and recrystallized from Water. Q-{HLQOONHOEECOS HZNCH2GOOH There is thus obtained 0.11 part of N-benzoylaminoacetyl-aminoacetic acid of melting point 206 C., that is COOH to say a yield of .46 percent, according to the reaction Q-GHZOCONHO'HiOONHCHmO0H+ represented by'the equation: SH

CONHOH2O O-SOHnC0 0H+ HZNCH2*COOH Q-O ONH'CH2C O-NHOHa-O OOH HS-OH;GO OH Example 4 Example 7 0.28 part of S-(carbobenzoxyamino-acetyl)-mercapto- 0.96 part of N-aminoacetyl-aminoacetic acid (glycylacetic acid (prepared according to Example 1 in applicaglycin) and 2.5 parts of S-(carbobenzoxyaminoacetyl)- tion Ser. No. 391,738, filed November 12, 1953) and mercaptoacetic acid are mixed and 4.0 parts by volume 0.075 part of aminoacetic acid are mixed With 1 part by of 4 N-caustic soda solution are added. After keeping volume of 2 N-caustic soda solution and the solution kept the solutionfor 20 hours at C. the pH value of 6.5 at 35 C. for 15 hours. After theaddition of 1 .part by is corrected again to 8.5 with 4 N-caustic soda solution. volume of 2 N-hydrochloric acid, the reaction mass is After a further 5 hours at 35 C. the Whole is acidified evaporated to dryness and unchanged S-(carbobenzoxy- 3r with concentrated hydrochloric acid to pH=2 and workamino-acetyl)mercapt0acetic acid, and also carbobenz- 0 ing up continued as described in Example 5. Thereis oxyamino-acetic acid, removed by means of alcohol. a yield of 1.8 parts or 79% of N-[N-(carbobenzoxy- Thus, by the reaction according to the equation Q-cm-o-c o-N11 crn-c os-om ooo:a ran-011200011 there is obtained N-carbobenzoxyaminoacetyl-aminoamino acetyl) --amino acetyl] aminoacetic acid (caracetic acid which, after crystallization from 2 parts by bobenzoxydiglycyl-glycin) melting at 204 C.

@omooounomcosnmoooa mNomcoNncmoooa Q-orno CON-HCHaO ON'HCHzC oNHorno 0 OH Hsomc OOH volume of water, melts at 178C. The yield is 0.19 part Example or -:percent of:tl1e-ca.lculated yield. 034 partiofss (carbgbgnzcxyamiflo acetyl) 111lople 5 salicylic acid and 0.13 p of N-aminQa s/Lamin 0.28 part of S-(carbobenzoxyamino-acetyl)-1nercaptoaceticacid are ,rcacted in an-analogous manner tha acetic acid and 0.075part of aminoacetic acid are heated describedin Example 6. There is, obtained-52% of the with 0.5 part by volume of -4'-N-caustic sodasolution for 55 calculated yield .of N-[N-(carbobenzoxy-aminoacetyl)- one hour at -85" -C. After cooling, the mixture is aminoacetyll-amino-acetic acid.

COOH

Q-cmooonnomoos +mNomooNncmooou COOH -CH2OCONHCH3C ONHCHaCONHGHsCOOH acidified with concentrated hydrochloric acid and cooled Examl 9 111 ice. T e pr c p ystals are separated fro 1.0 partof DL-alanylglycin and 2.3 parts of S-(,c,arthe mother hquor, Washed with cold water and dried. 7O bobenzoxyamino a etyl) mercapto acetic acid are After drying, soluble compounds are dissolved out with mixed and 3 75 parts b volume of 4 N-caustic d s l absolute 211601101 and the residue ystallized fI m tion are added. The solution is kept for 24 hours at 35 water. Yield: 0.180 part of N-carbobenzoxyamino- C, d then acidified with 8.5 parts by volume of 2 acetyl-aminoacetic acid melting at 178 C., correspond- N-hydrochloric acid and then saturated with sodium chloing to 67% of'the calculated yield. 7 7 ride. The oily reaction product is extracted with ethyl 6 acetate and crystallized with a mixture-of ethyl acetate promoter) and kept at 35 C. for 17 hours. The soluand ether. Yield: 1.68 parts=73% of carbobenzoxytion is given a pH of 7 by means of dilute caustic soda amino-acetyl-Dbalanyl-glycin melting at 150 C. Resolution, and the reaction product is then filtered oil and crystallization from water does not change the properrecrystallized from water. it melts at 25 l-253 C. Acties. After drying at 10- pressure of mercury the comcording'to the analysis the substance is N-(benzoylarninopound is analytically pure. acetyl)-N'-isonicotinyl-hydrazine.

CH3 @CHEOCONHGHQC osonioooH H:N(JHCONHCH|COOH l @0390 C ONHCHaO-ONHCHC ONHCHBC O OH HSCHaOOOH Example The following yields are obtained:

0.2 part of DzL-methionine and 0.4 part of S-(carbopH: Approx. percent benzoxyamino -acetyl) mercaptoacetic acid are mixed 2 -i 67 and 0.7 part by volume of 4 N-caustic soda solution is 3 100 added. After keeping the solution for hours at 4 91 C. the Whole is acidified with 1.5 parts by volume of 20 5 67 2 N-hydrochloric acid and extracted with ethyl acetate. 6

Q-oomromoosomooon HQNNHCO \r \\N -c ONHOlEbClONHNHCO- nsomo 0011 From the purified and dried extracts of ethyl acetate there 30 Example 13 can be obtained by concentration and addition of petro- 1.2 parts of S-(carbobenZoxyamino-acetyl)-mercaptoleum ether 0.22 part of N-(carbobenzoxyaminoacetic acid are dissolved in 10 parts by volume of water acetyl)-D:L-methionine melting at 123 C. and 5 parts by volume of l N-caustic soda solution and ({JHr-CHg-S-(LHs -omoconncmcoscmcooa mNcnooon OHr-CHa-S-CHa CHgOO ONHCHaCONHHC OOH HSCHaC O OH 0.13 part by volume of ethylene-diamine is added.

Crystallization occurs immediately. After 4 hours at a Example 11 temperature of 35 C. the crystallizate is filtered with suction and Washed with sodium carbonate solution, dilute 0.31 part of S (benzoylammo acetyl) h 0 a y C 45 hydrochloric acid and with water. Recrystallization is acid and 0.075 part Of amino-acetic acid reacted in the eflected from absolute alcohol, Melting point=2l2-213 same manner as described in Ex mp 6 yield P C. According to the analysis the substance is N:N'-diof N-(benzoylamino-acetyl)-aminoacetic acid (hipp ryl- (carbobenzoxyaminoacetyl) -ethylenediamine. The yield -omooo1-rnom0oscnooon HaNCHzCHaNHg znsomooon @omooomaomo O-NH @omooonrromo o-z rn Instead of the amines mentioned in the above examples the following reactants may be used: serine, cystine, C 0 OH aminobutyric acid, valine, leucine, phenylalanine, tyrosine,

tryptophan, histidine, 'y-carbobenzoxy-ornithine, aspara- ONHCHEC HENOHQCOOH gine, asparaginic acid, glutaminic acid, proline, diglycine, methionyl-glycine and glutathione. Furthermore there H000 may also be used other S-aminoacyl-mercaptan com- CONH 3H2OONH 3H2COOH+ pounds such as for example S-(phthalylamino-acetyl)- HS- mercaptoacetic acid, S-(benzoylamino-acetyl)-fi-mercaptopropionic acid, S-aminoacetyl-mercapto-acetic acid,

Example 1 S [N (N carbobenzoxyaminoacetyl amiuoacetyl) 0.28 part of S-(benzoylamino-acetyl)mercapto-acetic aminoacetyl]'mercaptoacefic "Acid or (N'amino' acid and 0.15 part of isonicotinic acid hydrazide are each acetyl aminoacfityl) aminoacetyl] mel'captoacetic dissolved in 20 parts by volume of acetate buffer of the acid, which are prepared according to the examples in desired pH (adding a little dimethylformamide as solution application Ser. No. 391,738, filed November 12, 1953,

glycin), i. e. in a yield of zyrsasu What is claimed is:

l. A process for aminoacylation, which comprises reacting a member selected from the group consisting of amines containing at least one hydrogen at the nitrogen atom and alcohols, with amino-thiolcarboxylic acid esters of the formula: v

wherein A represents an amino group, X stands for the divalent radical linking the amino and carboxyl group of a natural amino acid, Y stands for a member selected from the group consisting of lower alkylene and phenylene radicals and Z stands for the carboxyl group, said reaction being conducted in an aqueous medium.

2. A process in accordance with claim 1 wherein metal salts of the S-aminoacylmercaptans are employed.

3. A process in accordance with claim 1, wherein compounds of the formula are employed in which A stands for an acylamino group.

4. A process in accordance with claim 1, wherein compounds of the formula are employed in which A stands for a free amino group.

5. A process in accordance with claim 1, wherein compounds of the formula are employed in which X and Y are members which separate 'A from C0 and S from Z by 1 to 4 carbon atoms.

6. Process according to claim 1, wherein S-(benzoylamino-acetyl)-mercaptoacetic acid isused as starting material. v

7. Process according to claim 1, wherein S-(carbobenzoxyamino-aceteyl)-mercaptoacetic acid is used as starting material.

8. Process according to claim 1, wherein 'S-(carbobenzoxyamino-acetyl)-thiosalicylic acid is used as starting material.

9. Process according to claim 1, wherein S-(benzoylamino-acetyl)-thiosalicylic acid is used as starting material.

10. A process in accordance with claim 1, wherein the reaction is carried out at a pH from about 3 to about 9 inclusive.

References Cited in the tile of this patent UNITED STATES PATENTS 2,709,164 Wieland May 24, 1955 

1. A PROCESS FOR AMIONACYLATION, WHICH COMPRISES REACTING A MEMBER SELECTED FROM THE GROUP CONSISTING OF AMINES CONTAINING AT LEAST ONE HYDROGEN AT THE NITROGEN ATOM AND ALCOHOLS, WITH AMINO-THIOLCARBOXYLIC ACID ESTERS OF THE FORMULA: 